Asymptomatic infection with clade 2.3.4.4b highly pathogenic avian influenza A(H5N1) in carnivore pets, Italy, April 2023.

In April 2023, an outbreak of clade 2.3.4.4b highly pathogenic avian influenza A(H5N1) viruses carrying the T271A mammalian adaptive mutation in the PB2 protein was detected in a backyard poultry farm in Italy. Five domestic dogs and one cat living on the premises had seroconverted in the absence of clinical signs. Virological and serological monitoring of individuals exposed to the virus proved the absence of human transmission, however, asymptomatic influenza A(H5N1) infections in mammalian pets may have important public health implications.

Neutralizing antibody titers six months after Comirnaty vaccination: kinetics and comparison with SARS-CoV-2 immunoassays.

OBJECTIVES: mRNA vaccines, including Comirnaty (BNT162b2 mRNA, BioNTech-Pfizer), elicit high IgG and neutralizing antibody (NAb) responses after the second dose, but the progressive decrease in serum antibodies against SARS-CoV-2 following vaccination have raised questions concerning long-term immunity, decreased antibody levels being associated with breakthrough infections after vaccination, prompting the consideration of booster doses. METHODS: A total number of 189 Padua University-Hospital healthcare workers (HCW) who had received a second vaccine dose were asked to collect serum samples for determining Ab at 12 (t12) and 28 (t28) days, and 6 months (t6m) after their first Comirnaty/BNT162b2 inoculation. Ab titers were measured with plaque reduction neutralization test (PRNT), and three chemiluminescent immunoassays, targeting the receptor binding domain (RBD), the trimeric Spike protein (trimeric-S), and surrogate viral neutralization tests (sVNT). RESULTS: The median percentages (interquartile range) for decrease in antibodies values 6 months after the first dose were 86.8% (67.1-92.8%) for S-RBD IgG, 82% (58.6-89.3%) for trimeric-S, 70.4% (34.5-86.4%) for VNT-Nab, 75% (50-87.5%) for PRNT50 and 75% (50-93.7%) for PRNT90. At 6 months, neither PRNT titers nor VNT-Nab and S-RBD IgG bAb levels correlated with age (p=0.078) or gender (p=0.938), while they were correlated with previous infection (p<0.001). CONCLUSIONS: After 6 months, a method-independent reduction of around 90% in anti-SARS-CoV-2 antibodies was detected, while no significant differences were found between values of males and females aged between 24 and 65 years without compromised health status. Further efforts to improve analytical harmonization and standardization are needed.

Pulmonary fibrosis in a dog as a sequela of infection with Severe Acute Respiratory Syndrome Coronavirus 2? A case report.

BACKGROUND: Interstitial lung disease is a heterogeneous group of conditions characterized by severe radiographic changes and clinicopathological findings. However, in the vast majority of cases, the cause remains unknown. CASE DESCRIPTION: In the present study, we reported the clinical case of a 3 years old female Bull Terrier presented in October 2020 to the Advanced Diagnostic Imaging Department of the Turin Veterinary Teaching Hospital with a progressive pulmonary illness characterized by dyspnea, exercise intolerance, and a diffuse and severe pulmonary interstitial pattern at imaging investigations. Considering the clinical findings, the dog was included in a serological survey for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection in companion animals, showing positive results. Due to the further clinical worsening, the owners opted for euthanasia. At necroscopy, dog showed severe and chronic bronchopneumonia compatible with a Canine Idiopathic Pulmonary Fibrosis and with serological features linked to a SARS-CoV-2 infection. CONCLUSIONS: The comparison of these lesions with those reported in humans affected by Coronavirus Disease 2019 (COVID-19) supports the hypothesis that these findings may be attributable to the post-acute sequelae of SARS-CoV-2 infection in a dog with breed predisposition to Canine Idiopathic Pulmonary Fibrosis (CIPF), although direct evidence of SARS-CoV-2 by molecular or antigenic approaches remained unsolved.

Analytical and clinical performances of a SARS-CoV-2 S-RBD IgG assay: comparison with neutralization titers.

OBJECTIVES: SARS-CoV-2 serology presents an important role in several aspects of COVID-19 pandemic. Immunoassays performances have to be accurately evaluated and correlated with neutralizing antibodies. We investigated the analytical and clinical performances of a SARS-CoV-2 RBD IgG assay, automated on a high throughput platform, and the correlation of the antibodies (Ab) levels with the plaque reduction neutralization (PRNT50) Ab titers. METHODS: A series of 546 samples were evaluated by SARS-CoV-2 RBD IgG assay (Snibe diagnostics), including 171 negative and 168 positive SARS-CoV-2 subjects and a further group of 207 subjects of the COVID-19 family clusters follow-up cohort. RESULTS: Assay imprecision ranged from 3.98 to 12.18% being satisfactory at low and medium levels; linearity was excellent in all the measurement range. Considering specimens collected after 14 days post symptoms onset, overall sensitivity and specificity were 99.0 and 92.5%, respectively. A total of 281 leftover samples results of the PRNT50 test were available. An elevated correlation was obtained between the SARS-CoV-2 RBD IgG assay and the PRNT50 titer at univariate (ρ=0.689) and multivariate (ρ=0.712) analyses. CONCLUSIONS: SARS-CoV-2 S-RBD IgG assay shows satisfactory analytical and clinical performances, and a strong correlation with sera neutralizing activity.

Prevalence of SARS-CoV-2 RNA on inanimate surfaces: a systematic review and meta-analysis.

Coronavirus disease (COVID-19) is a respiratory disease affecting many people and able to be transmitted through direct and perhaps indirect contact. Direct contact transmission, mediated by aerosols or droplets, is widely demonstrated, whereas indirect transmission is only supported by collateral evidence such as virus persistence on inanimate surfaces and data from other similar viruses. The present systematic review aims to estimate SARS-CoV-2 prevalence on inanimate surfaces, identifying risk levels according to surface characteristics. Data were obtained from studies in published papers collected from two databases (PubMed and Embase) with the last search on 1 September 2020. Included studies had to be papers in English, had to deal with coronavirus and had to consider inanimate surfaces in real settings. Studies were coded according to our assessment of the risk that the investigated surfaces could be contaminated by SARS-CoV-2. A meta-analysis and a metaregression were carried out to quantify virus RNA prevalence and to identify important factors driving differences among studies. Thirty-nine out of forty retrieved paper reported studies carried out in healthcare settings on the prevalence of virus RNA, five studies carry out also analyses through cell culture and six tested the viability of isolated viruses. Overall prevalences of SARS-CoV-2 RNA on high-, medium- and low-risk surfaces were 0.22 (CI95 [0.152-0.296]), 0.04 (CI95 [0.007-0.090]), and 0.00 (CI95 [0.00-0.019]), respectively. The duration surfaces were exposed to virus sources (patients) was the main factor explaining differences in prevalence.

Functional characterization of a plant-produced infectious bursal disease virus antigen fused to the constant region of avian IgY immunoglobulins.

Infectious bursal disease virus (IBDV) is the cause of an economically important highly contagious disease of poultry, and vaccines are regarded as the most beneficial interventions for its prevention. In this study, plants were used to produce a recombinant chimeric IBDV antigen for the formulation of an innovative subunit vaccine. The fusion protein (PD-FcY) was designed to combine the immunodominant projection domain (PD) of the viral structural protein VP2 with the constant region of avian IgY (FcY), which was selected to enhance antigen uptake by avian immune cells. The gene construct encoding the fusion protein was transiently expressed in Nicotiana benthamiana plants and an extraction/purification protocol was set up, allowing to reduce the contamination by undesired plant compounds/proteins. Mass spectrometry analysis of the purified protein revealed that the glycosylation pattern of the FcY portion was similar to that observed in native IgY, while in vitro assays demonstrated the ability of PD-FcY to bind to the avian immunoglobulin receptor CHIR-AB1. Preliminary immunization studies proved that PD-FcY was able to induce the production of protective anti-IBDV-VP2 antibodies in chickens. In conclusion, the proposed fusion strategy holds promises for the development of innovative low-cost subunit vaccines for the prevention of avian viral diseases.

Isolation of coagulase-positive staphylococci from bitches' colostrum and milk and genetic typing of methicillin-resistant Staphylococcus pseudintermedius strains.

BACKGROUND: Among the coagulase-positive, potentially pathogenic staphylococci, Staphylococcus pseudintermedius has been frequently isolated from bitches' milk. This organism colonizes the mammary gland or causes infection, while S. aureus has been only occasionally reported. The objective of this study was to investigate the occurrence and persistence of coagulase-positive staphylococci in the colostrum and milk of postpartum bitches, either treated or untreated with antimicrobials, and to assess the incidence, antibiotic resistance profile and genetic type of the methicillin-resistant strains. On postpartum D1, D7 and D15, drops of secretion were collected from the mammary glands of 27 postpartum bitches, nine of which were treated with antimicrobials. Coagulase-positive staphylococci were identified, antimicrobial susceptibility and the presence of mecA were tested and the genetic profile of methicillin-resistant strains was assessed. RESULTS: Staphylococcus pseudintermedius was the only coagulase-positive staphylococcus isolated, and its presence was detected in 21 out of 27 bitches and in 66 out of 145 swabs. In a single bitch, it caused puerperal mastitis. In untreated bitches, the frequency of isolation was lower in colostrum than in milk. All of the isolates except one were resistant to at least three antimicrobial classes, while 14 out of 66 S. pseudintermedius strains were methicillin-resistant mecA positive (MRSP) and were isolated from eight bitches housed in the same breeding kennel. A significant association was found between antimicrobial treatment and the presence of MRSP. Six of the 12 typed isolates belonged to spa-type t02 carrying SCCmec II/III, and another six were non-typeable with spa carrying SCCmec IV. The t02-SCCmec II/III isolates were sequence type (ST) 71; four NT-SCCmec IV isolates were ST258 and two were ST369. PFGE showed that isolates from the same dog had identical band patterns, while isolates from different dogs had unique band patterns. MRSP strains showed multidrug resistance profiles. CONCLUSIONS: Our results confirm that S. pseudintermedius is the most frequently isolated coagulase-positive staphylococcus from bitches' milk. The isolation of several different strains of MRSP with different genetic characteristics in the same kennel and the fact that two of the strains belonged to a sequence type (ST) described for the first time are noteworthy findings.

H7N6 highly pathogenic avian influenza in Mozambique, 2023.

On 13 October 2023, the National Directorate for Livestock Development in Mozambique was notified of a suspected outbreak of avian influenza in commercial layers. Samples were screened by real-time and conventional RT-PCR and were positive for both H7 and N6. Full genome sequences were obtained for three representative samples. Sequence analysis of the H7 cleavage site confirmed that the viruses were highly pathogenic (i.e. 333- PEPPKGPRFRR/GLF-346). In addition, the H7 and N6 sequences were highly similar (from 99.4-99.5% and 99.6-99.7% for the HA gene and the NA gene, respectively) to the sequences of a H7N6 virus identified in the Republic of South Africa in May 2023 indicating a similar origin of the viruses. The identification of H7N6 HPAIV in Mozambique has important implications for disease management and food security in the region.

Vaccination of poultry against highly pathogenic avian influenza - Part 2. Surveillance and mitigation measures.

Selecting appropriate diagnostic methods that take account of the type of vaccine used is important when implementing a vaccination programme against highly pathogenic avian influenza (HPAI). If vaccination is effective, a decreased viral load is expected in the samples used for diagnosis, making molecular methods with high sensitivity the best choice. Although serological methods can be reasonably sensitive, they may produce results that are difficult to interpret. In addition to routine molecular monitoring, it is recommended to conduct viral isolation, genetic sequencing and phenotypic characterisation of any HPAI virus detected in vaccinated flocks to detect escape mutants early. Following emergency vaccination, various surveillance options based on virological testing of dead birds ('bucket sampling') at defined intervals were assessed to be effective for early detection of HPAIV and prove disease freedom in vaccinated populations. For ducks, virological or serological testing of live birds was assessed as an effective strategy. This surveillance could be also applied in the peri-vaccination zone on vaccinated establishments, while maintaining passive surveillance in unvaccinated chicken layers and turkeys, and weekly bucket sampling in unvaccinated ducks. To demonstrate disease freedom with > 99% confidence and to detect HPAI virus sufficiently early following preventive vaccination, monthly virological testing of all dead birds up to 15 per flock, coupled with passive surveillance in both vaccinated and unvaccinated flocks, is recommended. Reducing the sampling intervals increases the sensitivity of early detection up to 100%. To enable the safe movement of vaccinated poultry during emergency vaccination, laboratory examinations in the 72 h prior to the movement can be considered as a risk mitigation measure, in addition to clinical inspection; sampling results from existing surveillance activities carried out in these 72 h could be used. In this Opinion, several schemes are recommended to enable the safe movement of vaccinated poultry following preventive vaccination.

Host Response of Syrian Hamster to SARS-CoV-2 Infection including Differences with Humans and between Sexes.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has highlighted the importance of having proper tools and models to study the pathophysiology of emerging infectious diseases to test therapeutic protocols, assess changes in viral phenotypes, and evaluate the effects of viral evolution. This study provided a comprehensive characterization of the Syrian hamster (Mesocricetus auratus) as an animal model for SARS-CoV-2 infection using different approaches (description of clinical signs, viral load, receptor profiling, and host immune response) and targeting four different organs (lungs, intestine, brain, and PBMCs). Our data showed that both male and female hamsters were susceptible to the infection and developed a disease similar to the one observed in patients with COVID-19 that included moderate to severe pulmonary lesions, inflammation, and recruitment of the immune system in the lungs and at the systemic level. However, all animals recovered within 14 days without developing the severe pathology seen in humans, and none of them died. We found faint evidence for intestinal and neurological tropism associated with the absence of lesions and a minimal host response in intestines and brains, which highlighted another crucial difference with the multiorgan impairment of severe COVID-19. When comparing male and female hamsters, we observed that males sustained higher viral RNA shedding and replication in the lungs, suffered from more severe symptoms and histopathological lesions, and triggered higher pulmonary inflammation. Overall, these data confirmed the Syrian hamster as a suitable model for mild to moderate COVID-19 and reflected sex-related differences in the response against the virus observed in humans.

A novel array of real-time RT-PCR assays for the rapid pathotyping of type I avian paramyxovirus (APMV-1).

Newcastle disease (ND) caused by virulent avian paramyxovirus type I (APMV-1) is a WOAH and EU listed disease affecting poultry worldwide. ND exhibits different clinical manifestations that may either be neurological, respiratory and/or gastrointestinal, accompanied by high mortality. In contrast, mild or subclinical forms are generally caused by lentogenic APMV-1 and are not subject to notification. The rapid discrimination of virulent and avirulent viruses is paramount to limit the spread of virulent APMV-1. The appropriateness of molecular methods for APMV-1 pathotyping is often hampered by the high genetic variability of these viruses that affects sensitivity and inclusivity. This work presents a new array of real-time RT-PCR (RT-qPCR) assays that enable the identification of virulent and avirulent viruses in dual mode, i.e., through pathotype-specific probes and subsequent Sanger sequencing of the amplification product. Validation was performed according to the WOAH recommendations. Performance indicators on sensitivity, specificity, repeatability and reproducibility yielded favourable results. Reproducibility highlighted the need for assays optimization whenever major changes are made to the procedure. Overall, the new RT-qPCRs showed its ability to detect and pathotype all tested APMV-1 genotypes and its suitability for routine use in clinical samples.

SARS-CoV-2 in Animal Companions: A Serosurvey in Three Regions of Southern Italy.

Several animal species have been found to be susceptible to SARS-CoV-2 infection. The occurrence of infection in dogs and cats living in close contact with owners deserves particular attention from public health authorities in a One Health approach. In this study, we conducted serological screening to identify SARS-CoV-2 exposure in the sera from dogs and cats in three regions of southern Italy sampled during the years 2021 and 2022. We collected 100 serum samples in 2021 (89 from dogs and 11 from cats) and 640 in 2022 (577 from dogs and 63 from cats). Overall, the ELISA positivity rate was found to be 2.7% (20/740), with higher seroprevalence in dogs. Serum neutralization tests confirmed positivity only in two samples collected from dogs, and the assays, performed with serologically distinct SARS-CoV-2 variants, showed variant-specific positivity. This paper shows that monitoring SARS-CoV-2 exposure in animals might be affected by the viral antigenic evolution, which requires continuous updates to the serological tests used. Serological surveys are useful in understanding the true extent of exposure occurring in specific animal populations, not suffering the same limitations as molecular tests, and could help in identifying the infecting virus if tests able to characterize the immune response are used. The use of variant-specific validated serological methods should always be considered in serosurvey studies in order to determine the real impact of emerging variants on animal populations and its implications for veterinary and human health, as well as to identify potential reservoirs of the virus and its evolutionary changes.

Vaccination of poultry against highly pathogenic avian influenza - part 1. Available vaccines and vaccination strategies.

Several vaccines have been developed against highly pathogenic avian influenza (HPAI), mostly inactivated whole-virus vaccines for chickens. In the EU, one vaccine is authorised in chickens but is not fully efficacious to stop transmission, highlighting the need for vaccines tailored to diverse poultry species and production types. Off-label use of vaccines is possible, but effectiveness varies. Vaccines are usually injectable, a time-consuming process. Mass-application vaccines outside hatcheries remain rare. First vaccination varies from in-ovo to 6 weeks of age. Data about immunity onset and duration in the target species are often unavailable, despite being key for effective planning. Minimising antigenic distance between vaccines and field strains is essential, requiring rapid updates of vaccines to match circulating strains. Generating harmonised vaccine efficacy data showing vaccine ability to reduce transmission is crucial and this ability should be also assessed in field trials. Planning vaccination requires selecting the most adequate vaccine type and vaccination scheme. Emergency protective vaccination is limited to vaccines that are not restricted by species, age or pre-existing vector-immunity, while preventive vaccination should prioritise achieving the highest protection, especially for the most susceptible species in high-risk transmission areas. Model simulations in France, Italy and The Netherlands revealed that (i) duck and turkey farms are more infectious than chickens, (ii) depopulating infected farms only showed limitations in controlling disease spread, while 1-km ring-culling performed better than or similar to emergency preventive ring-vaccination scenarios, although with the highest number of depopulated farms, (iii) preventive vaccination of the most susceptible species in high-risk transmission areas was the best option to minimise the outbreaks' number and duration, (iv) during outbreaks in such areas, emergency protective vaccination in a 3-km radius was more effective than 1- and 10-km radius. Vaccine efficacy should be monitored and complement other surveillance and preventive efforts.

Detailed epitope mapping of SARS-CoV-2 nucleoprotein reveals specific immunoresponse in cats and dogs housed with COVID-19 patients.

Since the initial emergence in December 2019, the novel Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has been reported in over 200 countries, representing an unprecedented challenge related to disease control worldwide. In this context, cases of human to animal transmission have been reported, raising concern about the potential role of companion animals in the pandemic and stressing the need for reliable animal testing. In the study, a detailed epitope mapping of SARS-CoV-2 nucleoprotein, using both human and pet sera, allowed the identification of the most antigenic region in the C-terminus domain of the protein, which was used to develop an experimental double antigen-based ELISA. A panel of pre-pandemic sera and sera of animals immunized against (or naturally infected with) related coronaviruses was used to assess assay specificity at 99.5%. Positive sera belonging to animals housed with COVID-19 patients were confirmed with the experimental double-antigen ELISA using Plaque Reduction Neutralization test (PRNT) test as gold standard. The availability of a serological assay that targets a highly specific viral antigen represents a valuable tool for multispecies monitoring of Coronavirus Disease 2019 (COVID-19) infection in susceptible animals.

Protective Efficacy of H9N2 Avian Influenza Vaccines Inactivated by Ionizing Radiation Methods Administered by the Parenteral or Mucosal Routes.

H9N2 viruses have become, over the last 20 years, one of the most diffused poultry pathogens and have reached a level of endemicity in several countries. Attempts to control the spread and reduce the circulation of H9N2 have relied mainly on vaccination in endemic countries. However, the high level of adaptation to poultry, testified by low minimum infectious doses, replication to high titers, and high transmissibility, has severely hampered the results of vaccination campaigns. Commercially available vaccines have demonstrated high efficacy in protecting against clinical disease, but variable results have also been observed in reducing the level of replication and viral shedding in domestic poultry species. Antigenic drift and increased chances of zoonotic infections are the results of incomplete protection offered by the currently available vaccines, of which the vast majority are based on formalin-inactivated whole virus antigens. In our work, we evaluated experimental vaccines based on an H9N2 virus, inactivated by irradiation treatment, in reducing viral shedding upon different challenge doses and compared their efficacy with formalin-inactivated vaccines. Moreover, we evaluated mucosal delivery of inactivated antigens as an alternative route to subcutaneous and intramuscular vaccination. The results showed complete protection and prevention of replication in subcutaneously vaccinated Specific Pathogen Free White Leghorn chickens at low-to-intermediate challenge doses but a limited reduction of shedding at a high challenge dose. Mucosally vaccinated chickens showed a more variable response to experimental infection at all tested challenge doses and the main effect of vaccination attained the reduction of infected birds in the early phase of infection. Concerning mucosal vaccination, the irradiated vaccine was the only one affording complete protection from infection at the lowest challenge dose. Vaccine formulations based on H9N2 inactivated by irradiation demonstrated a potential for better performances than vaccines based on the formalin-inactivated antigen in terms of reduction of shedding and prevention of infection.

One-Year Surveillance of SARS-CoV-2 Exposure in Stray Cats and Kennel Dogs from Northeastern Italy.

Dogs and cats are susceptible to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). During the pandemic, several studies have been performed on owned cats and dogs, whereas limited data are available on the exposure to stray animals. The objective of this study was to investigate the exposure to SARS-CoV-2 of feral cats and kennel dogs in northeastern Italy, through serological and molecular methods. From May 2021 to September 2022, public health veterinary services collected serum, oropharyngeal, and rectal swab samples from 257 free-roaming dogs newly introduced to shelters, and from 389 feral cats examined during the routinely trap-neutered-return programs. The swabs were analyzed for viral RNA through a real-time reverse transcriptase PCR (rRT-PCR), and sera were tested for the presence of the specific antibody against SARS-CoV-2 (enzyme-linked immunosorbent assay). Serology was positive in nine dogs (9/257) and three cats (3/389), while two asymptomatic cats tested positive to rRT-PCR. One cat turned out to be positive both for serology and molecular analysis. In addition, this study described the case of a possible human-to-animal SARS-CoV-2 transmission in a cat that travelled in close contact to a COVID-19-positive refugee from Ukraine. This study shows that SARS-CoV-2 can infect, in natural conditions, stray cats and kennel dogs in northeastern Italy, although with a low prevalence.

Emergence of a Reassortant 2.3.4.4b Highly Pathogenic H5N1 Avian Influenza Virus Containing H9N2 PA Gene in Burkina Faso, West Africa, in 2021.

Since 2006, the poultry population in Burkina Faso has been seriously hit by different waves of Highly Pathogenic Avian Influenza (HPAI) H5N1 epizootics. In December 2021, three distinct regions of Burkina Faso, namely, Gomboussougou, Bonyollo, and Koubri, detected HPAI H5N1 viruses in poultry. Whole genome characterization and statistical phylogenetic approaches were applied to shed light on the potential origin of these viruses and estimate the time of virus emergence. Our results revealed that the HPAI H5N1 viruses reported in the three affected regions of Burkina Faso cluster together within clade 2.3.4.4b, and are closely related to HPAI H5N1 viruses identified in Nigeria and Niger in the period 2021-2022, except for the PA gene, which clusters with H9N2 viruses of the zoonotic G1 lineage collected in West Africa between 2017 and 2020. These reassortant viruses possess several mutations that may be associated with an increased zoonotic potential. Although it is difficult to ascertain where and when the reassortment event occurred, the emergence of a H5N1/H9N2 reassortant virus in a vulnerable region, such as West Africa, raises concerns about its possible impact on animal and human health. These findings also highlight the risk that West Africa may become a new hotspot for the emergence of new genotypes of HPAI viruses.

Rapid spread of a new West Nile virus lineage 1 associated with increased risk of neuroinvasive disease during a large outbreak in northern Italy, 2022: One Health analysis.

BACKGROUND: A new strain of WNV lineage 1 (WNV - 1) emerged in the Veneto Region, northern Italy, in 2021, eight years after the last outbreak of WNV - 1 in Italy. The virus, which co-circulates with WNV-2, has become endemic in the Region, where, in 2022, most human cases of neuroinvasive disease (WNND) reported in Europe have occurred. METHODS: Comparative analysis of the epidemiology and clinical presentation of WNV-1 and WNV-2 infection in humans, as well as the temporal and geographic distribution of WNV-1 and WNV-2 among wild birds and Culex pipiens mosquitoes in Veneto, from May 16th to August 21st, 2022, to determine if the high number of WNND cases was associated with WNV-1. RESULTS: As of August 21st, 2022, 222 human cases of WNV infection were confirmed by molecular testing, including 103 with fever (WNF) and 119 with WNND. WNV lineage was determined in 201 (90.5%) cases, comprising 138 WNV-1 and 63 WNV-2 infections. During the same period, 35 blood donors tested positive, including 30 in whom WNV lineage was determined (13 WNV-1 and 17 WNV-2). Comparative analysis of the distribution of WNV-1 and WNV-2 infections among WNND cases, WNF cases and WNV-positive blood donors showed that patients with WNND were more likely to have WNV-1 infection than blood donors (odds ratio 3.44; 95% CI 95% 1.54 to 8.24; p = 0.0043). As observed in humans, in wild birds WNV-1 had higher infectious rate (IR) and showed a more rapid expansion than WNV-2. At variance, the distribution of the two lineages was more even in mosquitoes, but with a trend of rapid increase of WNV-1 IR over WNV-2. CONCLUSIONS: Comparative analysis of WNV-1 vs WNV-2 infection in humans, wild birds, and mosquitos showed a rapid expansion of WNV-1 and suggested that WNV-1 infected patients might have an increased risk to develop severe disease.

Molecular Monitoring of SARS-CoV-2 in Different Sewage Plants in Venice and the Implications for Genetic Surveillance.

Wastewater-based epidemiology is now widely used as an indirect tool to monitor the spread of SARS-CoV-2. In this study, five different sample matrices representing diverse phases of the wastewater treatment process were collected during the second wave of SARS-CoV-2 from two wastewater treatment plants (WWTPs) serving the Civil Hospital and Sacca Fisola island in Venice, Italy. Positive SARS-CoV-2 detections occurred at both WWTPs, and data on viral genome detection rate and quantification suggest that the pellet (i.e., the particulate resulting from the influent) is a sensitive matrix that permits reliable assessment of infection prevalence while reducing time to results. On the contrary, analysis of post-treatment matrices provides evidence of the decontamination efficacy of both WWTPs. Finally, direct sequencing of wastewater samples enabled us to identify B.1.177 and B.1.160 as the prevalent SARS-CoV-2 lineages circulating in Venice at the time of sampling. This study confirmed the suitability of wastewater testing for studying SARS-CoV-2 circulation and established a simplified workflow for the prompt detection and characterization of the virus.

A systematic review of human coronaviruses survival on environmental surfaces.

The current pandemic caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has led people to implement preventive measures, including surface disinfection and use of alcohol-based hand gel, in order to avoid viral transmission via fomites. However, the role of surface transmission is still debated. The present systematic review aims to summarize all the evidence on surface survival of coronaviruses infecting humans. The analysis of 18 studies showed the longest coronavirus survival time is 28 days at room temperature (RT) on different surfaces: polymer banknotes, vinyl, steel, glass, and paper banknotes. Concerning SARS-CoV-2 human infection from contaminated surfaces, dangerous viral load on surfaces for up to 21 days was determined on polymer banknotes, steel, glass and paper banknotes. For viruses other than SARS-CoV-2, the longest period of survival was 14 days, recorded on glass. Environmental conditions can affect virus survival, and indeed, low temperatures and low humidity support prolonged survival of viruses on contaminated surfaces independently of surface type. Furthermore, it has been shown that exposure to sunlight significantly reduces the risk of surface transmission. Although studies are increasingly investigating the topic of coronavirus survival, it is difficult to compare them, given the methodology differences. For this reason, it is advisable to define a reference working protocol for virus survival trials, but, as an immediate measure, there is also a need for further investigations of coronavirus survival on surfaces.